Space flight has profound effects on the immune system of humans, monkeys and rodents. Several factors including microgravity, lack of load-bearing, stress, acceleration forces, and irradiation have been proposed to contribute to the changes of the immune system. However, the exact mechanisms by which these factors affect the immune system remain to be established. Simulation of these factors with ground animal models (e.g., hindlimb suspension in rodents) has been used to replicate the effects of space flight on immune system. Our recent studies have shown that opioids could induce Fas expression. In addition, restraint stress in mice induces Fas-mediated apoptosis in splenocytes in an endogenous opioid-dependent manner. Since space flight conditions resemble both physical and psychological stress to humans and animals, we hypothesize that immunosuppression during space flight is a result of the expression of Fas induced by increased production of endogenous opioids. We propose the following aims to test this hypothesis: 1) Determine the role of endogenous opioids in lymphocyte reduction induced by hindlimb suspension. At different times after suspension, mice will be treated with opioid receptor antagonists. Lymphocytes will be isolated from peripheral blood, spleen, and lymph nodes, and assayed for lymphocyte number, responsiveness to mitogenic stimulation, and the production of cytokines (INF-gamma, TNF, and IL-2) and antibodies. The number of apoptotic cells in the spleen and lymph nodes will be determined. 2) Since endogenous opioid production during space flight has not been studied, we will determine the level of endogenous opioids such as beta-endorphins, dynorphin, and enkephalin at different times after subjection to hindlimb suspension. 3) We will determine whether administration of anti-Fas ligand antibody or soluble recombinant Fas protein could block hindlimb suspension-induced lymphocyte reduction. In addition, the expression of Fas and FasL in animals subjected to hindlimb suspension will also be examined by northern blot analysis and surface staining. Furthermore, mice with Fas or FasL mutation will be used to verify the role of Fas in this process. The overall goal of these studies is to determine the mechanism(s) underlying immunosuppression associated with ground simulation of space flight, via endogenous opioids-modulated lymphocyte apoptosis. The results should lead to an understanding of the effects of space flight environment on human immune system.